Liver cell therapies: cellular sources and grafting strategies / 医学前沿

The liver has a complex cellular composition and a remarkable regenerative capacity. The primary cell types in the liver are two parenchymal cell populations, hepatocytes and cholangiocytes, that perform most of the functions of the liver and that are helped through interactions with non-parenchymal cell types comprising stellate cells, endothelia and various hemopoietic cell populations. The regulation of the cells in the liver is mediated by an insoluble complex of proteins and carbohydrates, the extracellular matrix, working synergistically with soluble paracrine and systemic signals. In recent years, with the rapid development of genetic sequencing technologies, research on the liver's cellular composition and its regulatory mechanisms during various conditions has been extensively explored. Meanwhile breakthroughs in strategies for cell transplantation are enabling a future in which there can be a rescue of patients with end-stage liver diseases, offering potential solutions to the chronic shortage of livers and alternatives to liver transplantation. This review will focus on the cellular mechanisms of liver homeostasis and how to select ideal sources of cells to be transplanted to achieve liver regeneration and repair. Recent advances are summarized for promoting the treatment of end-stage liver diseases by forms of cell transplantation that now include grafting strategies.

Continued administration of antithrombotic agents during transperineal prostate biopsy

Purpose To determine the safety of continued administration of antithrombotic agents during transperineal (TP) prostate biopsy. Patients and Methods A total of 811 men who underwent transrectal ultrasound (TRUS)-guided TP biopsy from January 2008 to June 2012 at our two institutions were retrospectively analyzed. Among these 811 men, 672 received no antithrombotic agents (group I), 103 received and continued administration of antithrombotic agents (group II), and 36 interrupted administration of antithrombotic agents (group III). Overall complications were graded and hemorrhagic complications were compared (group I with group II) using propensity score matching (PSM) analysis. Results An overall complication rate of 4.6% was recorded. Hemorrhagic complications occurred in 1.8% and they were virtually identical in all the three groups, and no severe hemorrhagic complications occurred. One patient in group III required intensive care unit admission for cerebral infarction. PSM analysis revealed no statistical difference between groups I and II with regard to the incidence of gross hematuria, perineal hematoma, and rectal bleeding. Multiple regression analysis revealed that hemorrhagic complications were associated with lower body mass index (<21 kg/m2, P=0.0058), but not with administration of antithrombotic agents. Conclusions Continued administration of antithrombotic agents does not increase the risk of hemorrhagic complications; these agents are well tolerated during TP biopsy. .

Freshly isolated hepatocyte transplantation in acetaminophen-induced hepatotoxicity model in rats / Transplante de hepatócitos recém-isolados em um modelo de hepatotoxicidade induzida por acetaminofeno em ratos

CONTEXT: Hepatocyte transplantation is an attractive therapeutic modality for liver disease as an alternative for orthotopic liver transplantation. OBJECTIVE: The aim of the current study was to investigate the feasibility of freshly isolated rat hepatocyte transplantation in acetaminophen-induced hepatotoxicity model. METHODS: Hepatocytes were isolated from male Wistar rats and transplanted 24 hours after acetaminophen administration in female recipients. Female rats received either 1x10(7) hepatocytes or phosphate buffered saline through the portal vein or into the spleen and were sacrificed after 48 hours. RESULTS: Alanine aminotransferase levels measured within the experiment did not differ between groups at any time point. Molecular analysis and histology showed presence of hepatocytes in liver of transplanted animals injected either through portal vein or spleen. CONCLUSION: These data demonstrate the feasibility and efficacy of hepatocyte transplantation in the liver or spleen in a mild acetaminophen-induced hepatotoxicity model.

CONTEXTO: O transplante de hepatócitos é uma modalidade terapêutica atrativa para doenças hepáticas como alternativa ao transplante hepático ortotópico. OBJETIVO: Investigar a factibilidade do uso de hepatócitos frescos isolados de ratos em um modelo de hepatotoxicidade induzida por paracetamol. MÉTODOS: Hepatócitos foram isolados de ratos Wistar machos e transplantados 24 horas após a administração de paracetamol em receptores fêmeas. As ratas receberam 1x10(7) hepatócitos ou tampão salina fosfato pela veia porta ou no baço e foram sacrificadas após 48 horas. RESULTADOS: Os níveis de alanina aminotransferase medidos durante o experimento não diferiram entre os grupos em nenhum momento. Análises moleculares e histológicas demonstraram a presença de hepatócitos no fígado dos animais transplantados pelo baço ou pela veia porta. CONCLUSÃO: Os dados indicam a factibilidade e eficácia do transplante de hepatócitos no fígado ou baço em um modelo de hepatotoxicidade leve induzida por paracetamol.

In vitro differentiation of human bone marrow mesenchy-mal stem cells into hepatocyte-like cells

Orthotropic liver transplantation [OLT] is the final procedure of both end stage and metabolic liver diseases. Hepatocyte transplantation is an alternative for OLT, but the sources of hepatocytes are limited. Bone marrow mesenchymal stem cells [BM-MSCs] can differentiate into hepatocyte-like cells and are a potential alternative source for hepatocytes. We aimed to investigate the differentiation potential of BM-MSCs into hepatocyte-like cells. Human BM-MSCs from a healthy donor were cultured and differentiated into hepatocyte-like cells. We investigated the expression of hepatocyte-specific markers in MSC-derived hepatocyte-like cells [MSC-HLC[s]] and evaluated their functionality using metabolic assays. MSC-HLCs expressed hepatocyte-specific markers at both mRNAand protein levels. In addition, the cells had the ability to uptake low density lipoprotein [LDL], clear ammonia, secrete albumin, and store glycogen. MSC-HLCs were transplanted into a familial hypercholesteromia patient. Human MSCs can be differentiated into partially functional hepatocyte-like cells. Thus, they could be a potential source for cell therapy in liver disorders

Peritoneal transplantation of human fetal hepatocytes for the treatment of acute fatty liver of pregnancy: a case report.

We report the case of a 26-year-old second gravida in the third trimester of pregnancy who presented with a history of nausea, repeated vomiting and jaundice. The patient was diagnosed as acute fatty liver of pregnancy. After delivery, the condition of the patient progressed to grade IV encephalopathy and did not improve despite all intensive clinical management measures. After 3 days in grade IV encephalopathy, the patient was infused 3 x 10(8) human foetal hepatocytes. The patient's level of consciousness started improving after 24 hours of foetal hepatocyte transfusion and she recovered completely within 7 days.

Cytokines' profile in fulminant hepatic failure: effect of hepatocyte transplantation

We used a previously described animal model of fulminant hepatic failure [FHF] to study the effect of hepatocyte transplantation [HcTx] on the cytokine profile. Three groups of Sprague Dawley rats were used. Group I [n=30] received intrasplenic HcTx. Group II [n=12] received intrasplenic injection of saline. Two days later, both groups underwent surgically induced FHF. Group III [n=30] underwent sham operation. Six animals of each group were euthanized preoperatively and on postoperative days 1, 7, 14 and 28 to study plasma cytokines; hepatocyte growth factor [HGF] and transforming growth factor beta one [TGF-beta 1]. Preoperatively, and after hepatocyte transplantation, Group I had a higher level of hepatocyte growth factor [HGF] than Groups II and III. On postoperative day 1, Group I had low levels of HGF and TGF-beta 1 and Group lI had the highest levels among the three groups. No animals in Group II survived beyond day 3 postoperatively. On day 14, HGF showed a rise in Group I as compared with Group III. HcTx has modified the level of cytokines in favor of liver regeneration in FHF rats

Liver gene therapy: prospects in the new century

Novel and exciting techniques have been developed for the genetic modification of hepatocytes. There are five broadly defined indications for direct gene transfer in liver therapy: 1] Gene replacement therapy, 2] Gene expression therapy, 3] Viral enzyme prodrug therapy, 4] Inhibition of gene expression, and 5] Repair of abnormal genes. The two main methods of gene delivery to the liver are: 1] viral vectors including Retroviruses, Adenoviruses, Adeno-associated, Simian virus 40, hybrid virus vectors, and 2] non-viral methods involving attachment of a therapeutic gene to a carrier. These may be either polymer based cationic carriers [conjugates] or lipid based vectors [liposomes]. The last decade of the second millennium has seen a lot of research work done in the field of genetics. Based on the many advances in the field, we hope that there is bright future for clinical applications of gene therapy for hepatic diseases

Hepatocyte isolation and transplantation in syngenic rats.

Refinement of techniques to isolate viable hepatocytes began in the late 1960's. It was established that perfusion of the intact liver as opposed to incubation of liver slices or chopped tissue increased the yield of cells. The present study aims to establish a simple, two-step, collagenase digestion method for hepatocyte isolation. A single inbred Fisher rat was used for hepatocyte isolation. The liver was perfused in-situ with perfusion buffer containing ethylene glycol bis N, N1, tetra acetic acid (EGTA), followed by the collagenase buffer. The liver was excised and gently minced. The tissue was resuspended in the collagenase buffer to complete dissociation. The cell suspension obtained was washed, centrifuged and filtered to complete the isolation procedure. The trypan blue exclusion test showed 80-85% cell viability. The isolated cells were transplanted into the splenic parenchyma of syngenic rats. Survival of the transplanted hepatocytes was confirmed by histological examination at the end of 90 days. This two step technique of in-situ liver perfusion gives a high yield of viable hepatocytes which show long term survival after transplantation.

Re-Engineering the liver with natural biomaterials

The extensive regenerative capacity of hepatocytes and the key roles of the liver in metabolic processes have generated interest in the liver as an appropriate target for cell and gene therapy. If cells were considered as natural biomaterials, then liver cell transplantation would fall within the general field of bioengineering. While unmodified hepatocytes engraft in the liver and ectopic sites, biological modifications and optimization of bioengineered systems would facilitate engraftment and survival of transplanted cells, especially in ectopic locations. Acute liver failure, chronic liver disease and metabolic deficiency states are among the conditions that can potentially be treated by cell transplantation. In acute liver failure, cell transplantation into the liver, along with the creation of an extrahepatic reservoir of cells might be required because engraftment and proliferation of transplanted cells in the liver needs time. In other situations, gradual liver repopulation alone might well be effective without additional manipulations.